This proposal aims to test an innovative new paradigm for the regulation of the sodium pump in the CNS. The sodium pump (Na,K-ATPase) consumes half of all the energy used in the brain. It is stably inhibited in ischemic tissue, which potentiates excitotoxicity. It contributes to neural and glial physiology through its regulation of Na and K and its electrogenic contribution to membrane potential. There is much evidence that it is actively regulated by transmitters and hormones, but the mechanisms remain frustratingly elusive at the molecular and biochemical level. We have recently shown that the Na,K-ATPase "gamma subunit" of the kidney acts as an accessory protein, modulating the Na,K-ATPase by reducing its affinity for Na and K. Others have shown that a homologue, CHIF, has a functionally opposite effect on kidney sodium pump properties, and the two are expressed in different nephron segments. These proteins are not expressed in the brain, but we have identified five other homologs that are. We will test the new hypothesis that they are all Na,K-ATPase regulators. The homologs (the FXYD gene family) share a conserved central membrane domain, but have divergent extracellular and intracellular domains. This predicts that they have different functional effects and are targeted by different regulatory mechanisms. In our preliminary data we show that one of the homologs is restricted to the cerebellar cortex molecular layer, and that it indeed associates with Na, K-ATPase. We will determine whether the other homologs have different cellular distributions, interact with different Na,K-ATPase isoforms, and have different functional effects on Na, K-ATPase properties. Two of them are new gene products that were first identified in our genomic studies, and pilot data are needed to make the transition from sequence to biology. The work promises to provide a significant breakthrough in the regulation of transport and energy metabolism in the brain, and may lead to neuroprotective therapy that could restore sodium pump activity in tissue-at-risk after stroke.